Effects of RU486 treatment after single prolonged stress depend on the post-stress interval.

The Single Prolonged Stress protocol is considered a model for PTSD, as it induces long lasting changes in rat behaviour and endocrine regulation. Previous work demonstrated that some of these changes can be prevented by treatment with the glucocorticoid receptor antagonist RU486, administered a week after the stressor. The current study evaluated the effects of an earlier intervention with RU486, as evaluated 1 week after SPS-exposure. Most RU486 effects occurred independent of prior stress, except for the reversal of a stress-induced increase in locomotor behaviour. The accompanying changes in gene expression depended on gene, brain region, and time. DNA methylation of the robustly down-regulated Fkbp5 gene was dissociated of changes in mRNA expression. The findings reinforce the long term effects of GR antagonist treatment, but also emphasize the need to evaluate changes over time to allow the identification of robust correlates between gene expression and behavioural/endocrine outcome of stressful experiences.

Expression of amygdala mineralocorticoid receptor and glucocorticoid receptor in the single-prolonged stress rats.

BACKGROUND: Post-traumatic stress disorder (PTSD) is an anxious disorder associated with low levels of corticosterone and enhanced negative feedback of the hypothalamic-pituitary-adrenal (HPA) axis. Previous studies showed that the amygdala not only has an excitatory effect on the HPA axis but also plays a key role in fear-related behaviors. Coticosterone exert actions through binding to the mineralocorticoid (MR) and glucocorticoid receptor (GR), which are abundant in the amygdala. In our previous study, down-regulation of MR and GR in the hippocampus of PTSD rats was found. But the roles of MR and GR in the amygdala of PTSD rats is incompletely understood. RESULTS: wistar rats were divided into 1 d, 7 d, 14 d groups after single prolonged stress (SPS) and control group. SPS is a reliable animal model of PTSD. Open field test (OF) and elevated plus maze tests (EPM) were performed to examine fear-related behaviors. Morphological changes of the ultrastructure of the amygdala neurons were assessed by transmission electron microscopy (TEM). Dual-immunofluorescence histochemistry was used to determined subcellular distribution and colocalization of MR- and GR-ir. Protein and mRNA of MR and GR was examined by western blotting and RT-PCR. OF and EPM showed enhanced fear in SPS rats. Abnormal neuronal morphology was discovered in the amygdala of SPS rats. The expression of MR- and GR-ir intensity, mRNA and protein within the amygdala decreased after SPS at 1 day, and then gradually recovered by 14 days, although the degree of decrease and recovery were different amongst techniques. We found no change in the MR/GR ratio at 3 levels of the amygdala. But more cytoplasmic distribution and decreased colocalization of MR- and GR-ir were observed in the amygdala after 7 days of SPS. CONCLUSION: These data suggest that change of MR and GR in the amygdala are involved in the mechanisms of fear in PTSD.

An Advanced Transcriptional Response to Corticosterone After Single Prolonged Stress in Male Rats.

Stress-related neuropsychiatric disorders are often accompanied by dysfunction of the hypothalamic-pituitary-adrenal (HPA) axis. In patients suffering from post-traumatic stress disorder (PTSD), increased sensitivity of glucocorticoid negative feedback has regularly been observed. The single prolonged stress (SPS) paradigm was developed to model increased negative feedback and other aspects of PTSD in rats. In this study, we used a setup that precluded the evaluation of negative feedback but rather served to test the hypothesis of the enhanced glucocorticoid receptor (GR) signaling in higher brain areas. We injected corticosterone or vehicle 7 days after SPS and evaluated plasma corticosterone, as well as gene expression in the dorsal hippocampus and amygdala. We observed a strikingly rapid change in the expression of established GR target genes (t = 30 min) only in the SPS group on exogenous corticosterone injection. Our results extend the notion of increased GR sensitivity in PTSD to include transcriptional responses in the hippocampus.

Late glucocorticoid receptor antagonism changes the outcome of adult life stress.

BACKGROUND: Stressors activate a wide spectrum of interacting hormonal and neuronal systems resulting in behavioral and physiological responses, with consequences for the development of psychopathology. Several recent studies demonstrated that treatment with the glucocorticoid receptor (GR) antagonist RU486 during adulthood normalized effects of early life stress. We aimed to evaluate the potential of RU486 to reverse stress-induced changes in an animal model of adult stress. METHOD: We employed the single-prolonged stress (SPS) model as a multimodal stress exposure protocol in male rats. SPS rats and unstressed controls were treated with RU486 on days 8, 9, 10 after stress exposure and the effects of treatment were evaluated after another 4 days. We determined body weight gain, corticosterone levels, behavioral reactivity in anxiety tests, and brain gene expression of c-fos, corticosteroid receptors, drivers of the stress response and genes (epi-)genitally linked to PTSD. RESULTS: RU486 affected body weight gain, corticosterone levels and open field behavior only in SPS rats. RU486 had history-independent effects in reducing fear in the elevated plus maze and fear conditioning behavior. Gene expression analysis showed a diversity of in- and interdependent effects of stress and RU486. CONCLUSION: The effects of RU486 applied 1 week after stress and measured 4 days after treatment demonstrate that in the state of post-SPS the GR-dependence of homeostatic processes has changed. This suggests that GR-mediated processes are part of allostatic regulation after adult stress. The normalization of a number of SPS-effects after RU486 treatment reinforces the potential of targeting GR for treatment of stress-related psychopathologies.

The role of ß-arrestin-2 on Fear/anxious-related memory in a rat model of Post-traumatic stress disorder.

BACKGROUND: Post-traumatic stress disorder (PTSD) can be categorised as a disorder of dysregulated fear processing. In the formation and development of PTSD, whether fear/anxious-related memory is regulated by ß-arrestin-2, and happened along the signal transduction pathways remains unknown. METHOD: We used single prolonged stress (SPS) as the animal model of PTSD. Next, elevated plus maze tests (EPM) was performed to examine fear/anxious memory- related behaviors. Then, we detected ß-arrestin-2, PDE-4, and signal transduction pathways with immunofluorescence, co-immunoprecipitation, immunohistochemistry, Elisa, western blot, RT-PCR, and real-time PCR. RESULTS: Our data indicated that SPS caused fear/anxious memory-related behaviors enhancement. The low expression of ß-arrestin-2, PDE-4 and their complex on SPS 7d, and high expression of signal transduction pathways on SPS7d in basolateral amygdala (BLA). CONCLUSIONS: That indicating that ß-arrestin-2 is critical for the formation of abnormal fear/anxious memory in PTSD; and fear/anxious memory occured through signal transduction pathways. Finally, these results suggest that ß-arrestin-2, PDE-4 and signal transduction pathways may be by influencing the fear/anxious memory thereby involved in the formation and development of PTSD.

Change of Rin1 and Stathmin in the Animal Model of Traumatic Stresses.

The molecular mechanism of fear memory is poorly understood. Therefore, the pathogenesis of post-traumatic stress disorder (PTSD), whose symptom presentation can enhance fear memory, remains largely unclear. Recent studies with knockout animals have reported that Rin1 and stathmin regulate fear memory. Rin1 inhibits acquisition and promotes memory extinction, whereas stathmin regulates innate and basal fear. The aim of our study was to examine changes in the expression of Rin1 and stathmin in different animal models of stress, particluarly traumatic stress. We used three animal traumatic stresses: single prolonged stress (SPS, which is a rodent model of PTSD), an immobilization-stress (IM) and a Loud sound stress (LSS), to examine the change and uniqueness in Rin1/stathmin expression. Behavioral tests of SPS rats demonstrated increased anxiety and contextual fear-conditioning. They showed decreased long-term potentiation (LTP), as well as decreased stathmin and increased Rin1 expression in the hippocampus and the amygdala. Expression of the stathmin effector, tubulin, and downstream molecules Rin1, Rab5, and Abl, appeared to increase. Rin1 and EphA4 were endogenously coexpressed in primary neurons after SPS stimulation. IM rats exhibited increased anxiety behavior and enhanced fear-conditioning to contextual and auditory stimuli. Similar changes in expression of Rin1/stathmin were observed in IM rats whereas no changes were observed in rats exposed to a loud sound. These data suggest that changes in expression of the Rin1 and stathmin genes may be involved in rodents with SPS and IM stresses, which provide valuable insight into fear memories under abnormal conditions, particularly in PTSD.

Single-prolonged stress induces apoptosis in the amygdala in a rat model of post-traumatic stress disorder.

OBJECTIVE: To detect the apoptosis-related Bax and Bcl-2 gene expression and apoptotic cell death in the amygdala region in the single-prolonged stress (SPS) rats. METHODS: A total of 100 male Wistar rats were randomly divided into a normal control group and SPS groups of 1d, 4d, 7d, and 14d. The expression of Bax and Bcl-2 was detected using immunohistochemistry and Western Blotting; TUNEL-staining and double-labeled flow cytometry (FCM) were employed for the detection and quantification of the apoptotic cells in the amygdala; morphological change of the subcellular structure in amygdala was observed by using the transmission electron microscopy (TEM). RESULTS: The ratio of Bax/Bcl-2 peaked at SPS 4d and then gradually decreased. The apoptosis peaked at SPS 4d. The TUNEL-positive cells were found in each SPS group and the TUNEL-positive cells rate peaked at SPS 4d. The morphological change of amygdala cells in each SPS group bears typical apoptotic characteristics. CONCLUSIONS: In the SPS rat brain, we found apoptotic process in the amygdala region which may relate to the pathogenesis of amygdala abnormal function in PTSD.

Ginsenoside Rb1 protects PC12 cells against ß-amyloid-induced cell injury.

Excessive accumulation of ß-amyloid (Aß) has been proposed as a pivotal event in the pathogenesis of Alzheimer's disease. Possible mechanisms underlying Aß-induced neuronal cytotoxicity include oxidative stress and apoptosis. Reactive oxygen species (ROS) have been proposed to be involved in the apoptotic mechanism of Aß-induced cytotoxicity. Ginsenoside Rb1 (GRb1), which is among the key compounds of ginsenoside, found in ginseng, may be a potent scavenger of ROS. To examine the potential protective effect of GRb1 in Aß25-35-induced cytotoxicity, cells were pre-treated with GRb1 for 24 h, and then Aß25-35 was added to the medium for an additional 24 h. Exposure to Aß led to the accumulation of ROS and lipid peroxidation, eventually causing a decrease in the Bcl-2/Bax ratio, caspase-3 activation, cell apoptosis and cell death. Pre-treatment with GRb1 not only inhibited Aß-induced ROS overproduction and lipid peroxidation, but also increased the Bcl-2/Bax ratio and attenuated caspase-3 activation, thereby improving cell survival. GRb1 may therefore act as a ROS scavenger, and such antioxidant properties may play a protective role against Aß-induced cell injury. Further exploration of GRb1 antioxidant properties may provide novel therapeutic strategies for the treatment of Alzheimer's disease.